Ramesh C. Khanal

Purified Blueberry Anthocyanins and Blueberry Juice Alter Development of Obesity in Mice Fed an Obesogenic High-Fat Diet†

Male C57BL/6J mice (25 days of age) were fed either a low-fat diet (10% kcal from fat) (LF) or a high-fat diet (45% kcal from fat) (HF45) for a period of 72 days. Blueberry juice or purified blueberry anthocyanins (0.2 or 1.0 mg/mL) in the drinking water were included in LF or HF45 treatments. Sucrose was added to the drinking water of one treatment to test if the sugars in blueberry juice would affect development of obesity. Total body weights (g) and body fat (%) were higher and body lean tissue (%) was lower in the HF45 fed mice compared to the LF fed mice after 72 days, but in mice fed HF45 diet plus blueberry juice or blueberry anthocyanins (0.2 mg/mL), body fat (%) was not different from those mice fed the LF diet. Anthocyanins (ACNs) decreased retroperitoneal and epididymal adipose tissue weights. Fasting serum glucose concentrations were higher in mice fed the HF45 diet. However, it was reduced to LF levels in mice fed the HF45 diet plus 0.2 mg of ACNs/mL in the drinking water, but not with blueberry juice. beta cell function (HOMA-BCF) score was lowered with HF45 feeding but returned to normal levels in mice fed the HF45 diet plus purified ACNs (0.2 mg/mL). Serum leptin was elevated in mice fed HF45 diet, and feeding either blueberry juice or purified ACNs (0.2 mg/mL) decreased serum leptin levels relative to HF45 control. Sucrose in drinking water, when consumption was restricted to the volume of juice consumed, produced lower serum leptin and insulin levels, leptin/fat, and retroperitoneal and total fat (% BW). Blueberry juice was not as effective as the low dose of anthocyanins in the drinking water in preventing obesity. Additional studies are needed to determine factors responsible for the differing responses of blueberry juice and whole blueberry in preventing the development of obesity.

Procyanidin content of grape seed and pomace, and total anthocyanin content of grape pomace as affected by extrusion processing.

UNLABELLED Grape juice processing by-products, grape seed and pomace are a rich source of procyanidins, compounds that may afford protection against chronic disease. This study was undertaken to identify optimal extrusion conditions to enhance the contents of monomers and dimers at the expense of large molecular weight procyanidin oligomers and polymers in grape seed and pomace. Extrusion variables, temperature (160, 170, and 180 degrees C in grape seed, and 160, 170, 180, and 190 degrees C in pomace) and screw speed (100, 150, and 200 rpm in both) were tested using mixtures of grape seed as well as pomace with decorticated white sorghum flour at a ratio of 30 : 70 and moisture content of 45%. Samples of grape seed and pomace were analyzed for procyanidin composition before and after extrusion, and total anthocyanins were determined in pomace. Additionally, chromatograms from diol and normal phase high-performance liquid chromatography were compared for the separation of procyanidins. Extrusion of both grape by-products increased the biologically important monomer and dimers considerably across all temperature and screw speeds. Highest monomer content resulted when extruded at a temperature of 170 degrees C and screw speed of 200 rpm, which were 120% and 80% higher than the unextruded grape seed and pomace, respectively. Increases in monomer and dimer contents were apparently the result of reduced polymer contents, which declined by 27% to 54%, or enhanced extraction facilitated by disruption of the food matrix during extrusion. Extrusion processing reduced total anthocyanins in pomace by 18% to 53%. PRACTICAL APPLICATIONS Extrusion processing can be used to increase procyanidin monomer and dimer contents in grape seed and pomace. Procyanidins in grape by-products have many health benefits, but most are present as large molecular weight compounds, which are poorly absorbed. Extrusion processing appears to be a promising technology to increase levels of the bioactive low molecular weight procyanidins.

Influence of extrusion processing on procyanidin composition and total anthocyanin contents of blueberry pomace.

Blueberry juice processing by-products are a rich source of procyanidins, which comprise a group of compounds shown to possess numerous health benefits, including protection against coronary heart disease, type II diabetes, and obesity. Most of the procyanidins present in blueberry pomace, however, are large molecular weight compounds that are poorly absorbed and show weak bioactivity compared to the smaller molecular weight monomers and dimers. The objective of our study was to identify optimal extrusion variables to enhance the contents of monomers and dimers at the expense of large molecular weight procyanidin oligomers and polymers. Extrusion variables temperature (160 and 180 degrees C) and screw speed (150 and 200 rpm) were tested using mixtures of blueberry pomace with decorticated white sorghum flour at a ratio of 30 : 70 and 45% moisture content. Extrudates were analyzed for procyanidin composition and total anthocyanin content. Extrusion of blueberry pomace increased the monomer, dimer, and trimer contents considerably at both temperature and screw speeds. The highest monomer content, obtained at 180 degrees C and 150 rpm screw speed, was 84% higher than the nonextruded control. Significantly higher levels of dimer and trimer contents were also obtained under these conditions. Increases in monomer, dimer, and trimer contents apparently were the result of reduced polymer contents, which was approximately 40% lower for samples extruded at 180 degrees C temperature and 150 rpm screw speed. Extrusion processing reduced total anthocyanin contents by 33% to 42% indicating that additional treatments are needed to retain the pigments. These results demonstrate that extrusion processing can be used to increase procyanidin monomer and dimers in blueberry pomace.

Urinary Excretion of Phenolic Acids in Rats Fed Cranberry

Dietary flavonoids can be converted into phenolic acids by colonic microflora. Phenolic acids can then be absorbed into the circulation and may contribute to the health-promoting effects of the parent compounds. Phenolic acids can be further metabolized in other tissues via methylation and conjugation with glucuronide or sulfate. The objectives of this study were to identify and quantify the urinary excretion of 19 phenolic acids and their conjugates in rats fed three levels of a concentrated cranberry powder (3.3, 6.6, and 33 mg/kg of diet). The basic diet used was AIN93G diet containing very low amounts of any polyphenolic compounds. Of the phenolic acids studied, the amounts excreted varied by 4 orders of magnitude, with hippuric acid being excreted in the highest quantities. Amounts of 4-hydroxyphenylacetic acid (4HPAA), 3-hydroxyphenylacetic acid (3HPAA), 3-hydroxyphenylpropionic acid (3HPPA), and 4-hydroxycinnamic acid (4HCA) excreted were in the range of 18-33 microg/mg creatinine in animals fed the highest level of cranberry powder, whereas phenylacetic acid (PAA), gallic acid (GA), 3,4-dihydroxyphenylacetic acid (34HPAA), 3,4-dihydroxybenzoic acid (34HBA), 3,4-dihydroxycinnamic acid (34HCA), and 4-hydroxy-3-methoxycinnamic acid (FA) were excreted in the urine in concentrations of 0.1-2 microg/mg creatinine. As the amount of cranberry in the diet was increased, the amount of 4HPAA excreted decreased but the percentage of conjugated 4HPAA excreted increased (from 57 to 91%). For other phenolic acids analyzed, the percentage excreted in the conjugated form was approximately constant across levels of cranberry in the diet and ranged from 65 to 100% for the individual phenolic acids. Studies of bioactivity and health effects need to consider more than just the compound(s) in the food, because they can be metabolized to other lower molecular weight compounds, which in turn may also be methylated or conjugated in some form that may affect the perceived health effects.

ENDOCRINE REGULATION OF CALCIUM TRANSPORT IN EPITHELIA

1 Calcium (re)absorption occurs in epithelia, including the intestine, kidney, mammary glands, placenta and gills (in the case of fish). 2 Calcium is transported across epithelia by two transport mechanisms, paracellular and transcellular, and the movement is regulated by a complex array of transport processes that are mediated by hormonal, developmental and physiological factors involving the gastrointestinal tract, bone, kidney and the parathyroids. 3 Clear understanding of the calcium transport pathways and their endocrine regulation is critical for minimizing various metabolic and health disorders at different physiological stages. Here, we first briefly review the calcium transport mechanisms before discussing in detail the endocrine factors that regulate calcium transport in the epithelia.

Dietary Black Raspberry Anthocyanins Do Not Alter Development of Obesity in Mice Fed an Obesogenic High-Fat Diet †

Anthocyanins (ACNs) from various foods have been shown to minimize the development of obesity in some animal models. The objective of the current study was to compare the effects of feeding purified black raspberry (BRB) ACNs or the freeze-dried whole BRB on the development of obesity. Male C57BL/6J mice (25 days of age) were assigned at random to treatments (7/treatment; 3/cage). The treatments included (1) control low-fat diet (10% calories from fat) (LF); (2) LF plus BRB juice in place of drinking water; (3) LF diet plus purified BRB ACNs in drinking water (1.25 mg/mL); (4) control high-fat diet (60% calories from fat) (HF60); (5) HF60 diet + BRB juice in place of drinking water; (6) HF60 diet + ACNs in drinking water (1.25 mg/mL); and (7) HF60 + freeze-dried whole BRB powder (21.7 g/kg of diet). Body weight gains in mice fed HF60 diet plus purified BRB ACNs tended to be lower after 56, 63, and 70 days than in mice fed HF60 alone. Body weights were increased at time of sacrifice, but heart, liver, and kidney weights as a percentage of body weight were decreased in mice fed HF60 diet compared to LF fed mice. Weights (g or g/body weight) of epididymal and retroperitoneal fat were increased in the HF60 fed mice compared to LF fed mice. Fasting serum glucose, leptin, and insulin levels as well as homeostasis assessment of insulin resistance (HOMA-IR) were elevated in mice fed the HF60 diet relative to LF-fed controls. Serum cholesterol, triglycerides, and monocyte chemoattractant protein-1 (MCP-1) were not altered by diet. Serum levels of resistin were increased in mice fed the HF60 diet compared to mice fed the LF diet. None of the responses measured were altered by whole BRB powder included in the diet relative to the HF60 control diet. Cyanidin containing di- or triglycosides in BRB was ineffective in altering the development of obesity in contrast to cyanidin-monoglycosides, which have been shown to be effective. The sugar moiety on the anthocyanidins may be an important factor in determining the response in the development of obesity.

Procyanidin Composition of Selected Fruits and Fruit Byproducts Is Affected by Extraction Method and Variety

Fruits and fruit byproducts are rich sources of polyphenols, including procyanidins, which are known to have numerous potential health benefits. This study investigated if varietal differences existed in the procyanidin composition of grape seed and if soaking in extraction solvent overnight prior to extraction improved the recovery of procyanidins from grape seed, grape pomace, and blueberry and cranberry powders. Riesling contained the highest amount of procyanidins, including the lower molecular weight monomers and dimers, followed by Chardonnay (60%), whereas Merlot contained much lower levels (14%) of individual and total procyanidins. A modified method of extraction whereby selected fruits and fruit byproducts were soaked in the extraction solvent overnight before the extraction process was begun increased procyanidins extracted by 24-100% from grape seeds and by 0-30% with berry procyanidin sources. The results indicate a wide variation in the procyanidin contents among different varieties of grape seeds that could have implications in the selection of procyanidin-rich germplasm. Soaking samples in the extraction solvent for 16 h resulted in increased procyanidins extracted and thus higher calculated concentrations in the food samples tested.

Effect of Dietary Blueberry Pomace on Selected Metabolic Factors Associated with High Fructose Feeding in Growing Sprague–Dawley Rats

An experiment was conducted to study the protective effect of feeding extruded and unextruded blueberry pomace (BBP) on selected metabolic parameters associated with metabolic syndrome in a model of high fructose (HF)-fed growing Sprague-Dawley rats. Treatments were as follows: (1) control (modified AIN-based diet); (2) HF diet (AIN diet with 58% fructose); (3) HF diet with 1.5% unextruded BBP; (4) HF diet with 1.5% extruded BBP; (5) HF diet with 3% unextruded BBP; and (6) HF diet with 3% extruded BBP. Compared with the control, HF feeding increased fasting plasma insulin and fasting and postprandial plasma triglycerides as well as homeostatic scores of insulin resistance and β-cell function, but not weight gain, diet intake and efficiency, abdominal fat, oral glucose tolerance, and fasting and postprandial plasma glucose, cholesterol, and leptin levels. Inclusion of unextruded or extruded BBP was effective in minimizing or ameliorating the fructose-induced metabolic anomalies, except postprandial plasma triglycerides, especially at 3% of the diet. In addition, unextruded or extruded BBP at 3% of the diet was also able to reduce plasma cholesterol and abdominal fat relative to the HF control, which may impart additional health benefits. Compared with the control, inclusion of unextruded or extruded BBP at both 1.5% and 3% resulted in lower total fat weight, and animals fed a diet supplemented with 3% unextruded BBP in fasting state or 3% unextruded BBP in fed state had lower leptin levels than the control. This is the first study demonstrating the beneficial effects of feeding blueberry pomace on health.

Urinary Excretion of Phenolic Acids in Rats Fed Cranberry, Blueberry, or Black Raspberry Powder

Dietary polyphenolics can be converted into smaller phenolic acids (PA) by microorganisms in the colon and may contribute to health benefits associated with the parent polyphenolics. Urinary excretion of 18 PA and their conjugates was studied, using HPLC-MS/MS, in rats fed AIN93G-based diets containing 5% (dry weight basis) of either cranberry (CB), blueberry (BB), or black raspberry (BRB). Hippuric, 4-hydroxyphenylacetic, 3-methoxy-4-hydroxyphenylacetic, and 4-hydroxybenzoic acids were excreted in greatest quantity in the urine over a 24 h period in all diets. Primary PA excreted in the berry diets were 4-hydroxycinnamic acid for CB; chlorogenic, ferulic, and 3,4-dihydroxycinnamic acids for BB; and 3-hydroxyphenylpropionic, 3-hydroxybenzoic, and 3-hydroxycinnamic acids for BRB. PA were present in conjugated form with cinnamic acid derivatives being 50-70% and phenylacetic acid derivatives conjugated <10%. Conjugated, and not just the free, PA are significant contributors to total urinary excretion.

Effect of Feeding Grape Pomace on Selected Metabolic Parameters Associated with High Fructose Feeding in Growing Sprague–Dawley Rats

The effect of feeding grape pomace on certain metabolic parameters associated with high fructose (HF) feeding was studied. Forty male growing Sprague-Dawley rats were randomly assigned into groups: (1) control; (2) HF; (3) HF with low-level (1.5% of diet) grape pomace (HF+LP), and (4) HF with high-level (5.0% of diet) grape pomace (HF+HP). The HF+LP and HF+HP diets provided 115 and 218 mg of procyanidins/kg, respectively. Compared with the controls, HF-fed animals consumed less and were smaller, whereas animals in the HF+LP and HF+HP groups were in between. A similar trend was observed for abdominal fat and abdominal fat as a percentage of body weight. No change in heart or kidney weight occurred. Liver weight as a percentage of body weight was higher for animals when fructose was included in the diet compared with those on control diet, and inclusion of grape pomace had no effect. Fasting plasma glucose, insulin, and triglyceride levels tended to be higher in animals fed HF diet, and grape pomace reduced their levels to values similar to the control animals. Compared with control animals, HF-fed animals had higher weekly postprandial plasma triglycerides, which were reduced by feeding grape pomace, but no change in plasma cholesterol was observed. Glucose intolerance was observed in animals fed HF diet and was accompanied by a 25% increase in homeostatic model assessment (HOMA) of insulin resistance. Inclusion of grape pomace increased glucose tolerance and insulin sensitivity. No significant change (P>.1) in HOMA of β-cell function or Quantitative Insulin-Sensitivity Check Index was observed. Overall, HF diet did not produce as strong a response of metabolic syndrome as has been shown in the literature. The inclusion of grape pomace in the diet was effective in modulating some aspects of metabolic parameters associated with metabolic syndrome, and the higher level of grape pomace in the diet produced a slightly better response than the lower level.